Many physiological processes in eukaryotic cells are regulated by the covalent modification of proteins by phosphorylation-dephosphorylation mechanisms. There is increasing evidence that these controls play primary regulatory roles in maintaining normal cellular metabolism during cell-growth and development as well as in altering metabolic processes in response to physiological and environmental stimuli (nutritional deficiency, anoxia, energy-depletion, elevated temperatures, hormones, mitogens, virus infection). Recent studies also implicate a role for protein phosphorylation in tumorigenicity and in the anti-viral effects of interferon. Many of these processes are regulated in part at the level of protein chain initiation. Protein synthesis in mammalian reticulocyte lysates is regulated by the phosphorylation of the Kappa-subunit of initiation factor eIF-2(eIF-2 Alpha). Phosphorylation of eIF-2 Alpha produces a potent inhibition of initiation in lysates. This phosphorylation is catalyzed by specific cyclic-nucleotide-independent eIF-2 Alpha kinases which are activated from latent inactive forms in response to at least three modes of physiological stress: (i) heme-deficiency; (ii) oxidized glutathione; (iii) double-stranded RNA. Similar eIF-2 Alpha kinases have been detected in other cells and tissues but their physiological function in translational control in these cells is not known. Reticulocytes also contain an eIF-2 Alpha phosphatase activity that reverses the effect of inhibitory kinases. The ratio of eIF-2 Alpha kinase: eIF-2 Alpha phosphatase activities regulates the extent of eIF-2 Alpha phosphorylation and hence the rate of protein synthesis initiation. The primary goal of the proposed study is to determine the physiological significance of the eIF-2 Alpha kinase-eIf-2 Alpha phosphatase system in the control of protein synthesis in reticulocytes and cultured mammalian cells. Specific research goals will focus on the (a) biochemical and molecular properties of eIF-2 Alpha kinases and eIF-2 Alpha phosphatases; (b) examination of their mechanism of regulation and expression; (c) function of this regulatory mechanism during normal cell-growth and development and in response to stress (nutritional deprivation, heat- and hypertonic-shock, anoxia). Regulation of translation by the phosphorylation of other initiation components (ribosomal proteins, other initiation factors and messenger ribonucleoproteins) will be examined in parallel.